Mutations in myosin VIIa (MYO7A) are the most common cause of Usher syndrome type 1. MYO7A is believed to be essential for the formation and function of the hair cell mechanotransduction (MET) tip link complex and the stereocilia ankle link, but its precise function in these complexes is not known. In preliminary studies, we discovered that the cochlea expresses multiple isoforms of MYO7A. In a genetically engineered mouse model in which the canonical isoform (Myo7a-C) is specifically deleted (Myo7a-?C mouse), MYO7A expression is severely diminished in inner hair cells (IHCs), and to a lesser degree in apical outer hair cells (OHCs). In contrast, deletion of the alternative isoform Myo7a-N (Myo7a-?N mouse) led to a significant reduction of MYO7A levels in OHCs, varying in intensity along the tonotopic axis. Analyses of these models led to the hypothesis that two major isoforms are expressed in a complementary manner in the cochlea: IHCs predominantly express the canonical isoform MYO7A-C, and much lower levels of the alternative isoform MYO7A-N. In OHCs, the two isoforms are expressed in opposing gradients along the tonotopic axis. This surprising feature of MYO7A expression gave rise to a novel conceptual framework and experimental tools to interrogate the functional role of MYO7A in the hair cell in the following three aims:
In Specific Aim (SA) 1, we propose to further investigate the variety of MYO7A isoforms, and their expression and localization in cochlear hair cells. To this end, we have already generated a mouse model in which one of the isoforms is genetically tagged, allowing us to determine its cellular and subcellular localization and characterize the isoform-specific interactome. In SA2, we will test the functional significance of each MYO7A isoform for hair cell MET and hearing performance. Preliminary electrophysiological studies show that genetic deletion of the canonical isoform affects resting open probability and current activation in response to fluid jet stimulation in IHCs, consistent with a putative role of MYO7A in tensioning the tip link complex. Finally, we ask why different types of hair cells express distinct isoforms of MYO7A. To address this, in SA3, we test the hypothesis that the differential expression of MYO7A isoforms serves to tune tip link tension in hair cells, thereby modulating MET current properties along the tonotopic axis. This project, through a multi-disciplinary collaboration that enabled the combination of molecular manipulations in the mouse, electrophysiology, imaging and biochemistry techniques, has the potential to provide critical insights into the function of an important deafness gene.
The mammalian auditory organ harbors two types of sensory cells called inner and outer hair cells. We made the unexpected and novel discovery that each cochlear cell type preferentially expresses a distinct isoform of the deafness gene Myosin VIIa (MYO7A). We hypothesize that intrinsic differences of the two MYO7A isoforms significantly influence the functional output of the two hair cell types. This project has the potential to reveal the molecular mechanisms underlying the functional specialization of different hair cell types, and will contribute fundamentally to our understanding of hearing and deafness.
